Erythropoiesis in the adult bone marrow is primarily homeostatic, producing a constant level of erythrocytes throughout adult life. This situation is dramatically different in the fetal liver during embryogenesis and in the adult spleen following acute erythroid stress. In both of these cases, erythropoiesis rapidly produces larger numbers of erythrocytes. Given that both the fetal liver and the spleen are sites of expansive erythropoiesis it has been suggested that the molecular mechanisms that regulate erythropoiesis in the fetal liver during development and the spleen during erythroid stress are similar, but distinct from steady state adult bone marrow erythropoiesis. This connection is evident in mice mutated at the flexed-tail (f) locus. f/f mutant mice exhibit a severe fetal anemia that resolves by 2 weeks after birth. Adult f/f mice exhibit normal steady state blood parameters, however, they exhibit a severe delay in the response to acute erythroid stress. We have identified a mutation in the Smad5 gene in f/f mice. Smad5 is a signaling protein-transcription factor that acts downstream of the BMP4 receptor. BMP4 plays a key role in the development of the development of hematopoietic cells and in particular the development of the erythroid lineage. This proposal outlines experiments designed to investigate the role of the BMP4/Smad5 signaling pathway in the expansive erythropoiesis characteristic of the fetal liver during development and the spleen during the response to erythropoietic stress. We will utilize the f/f mice as a means to dissect the role of BMP4 and Smad5 in hematopoesis and erythropoiesis. First, we determine the role of BMP4 and Smad5 in the expansion and differentiation of stress erythroid progenitors in the spleen following acute stress. Second we will analyze the role of BMP4 and Smad5 in the development of hematopoietic stem cells and multipotential cells in the Aorta- Gonad-Mesonephros (AGM) region of the embryo. Third, we will analyze the role of BMP4 and Smad5 in the development and differentiation of fetal liver erythroid progenitors. These analyses will provide important basic information that could be used to develop therapies for anemia and the treatment of traumatic blood loss.